Current combination antiretroviral therapy (ART) has been successful at prolonging life for patients with HIV. Since current ART regimens cannot eradicate the HIV-1 virus, patients remain chronically infected and must remain on ART indefinitely once the treatment threshold is met. Lifelong ART requires sustained daily adherence, and waning of adherence often leads to therapeutic failure and the emergence of resistant viruses. Multiple adherence measures and interventions have been studied to improve declining ART adherence, with limited success. Each strategy has inherent limitations and varying degrees of efficiency and cost. Laboratory monitoring of ART regimens consists primarily of measuring CD4 T-cell counts and plasma HIV-1 RNA levels. The goal of ART treatment is to achieve undetectable plasma HIV-1 RNA levels below 50 copies/mL, based on the limit of detection of current clinical assays. Rising HIV-1 viremia is used by clinicians as a marker of possible regimen failure, suboptimal adherence, and/or emergence of drug resistance. Establishing virologic failure requires confirmation of persistently increased HIV-1 RNA levels beyond a predetermined threshold. During this period, continued viral replication, evolution and emergence of ART resistance may occur, which limits current and future treatment options for patients. These limitations underscore the need for an early, objective and sensitive marker of loss of virologic suppression that could allow for interventions prior to therapeutic failure and the development of drug resistance. HIV-1 RNA levels below 50 copies/mL are readily detected by more sensitive assays. HIV-1 residual viremia averages between 1-3 copies/mL on suppressive ART regimens. Our pilot studies suggest that patients with eventual virologic failure develop increasing residual viremia levels 4 to 12 weeks prior to virologic rebound above 50 copies/mL. Earlier detection of viral replication provides an objective assessment of loss of virologic control, expanding the timeframe for clinical interventions to prevent regimen failure and emergence of drug resistance. For this project, we will collaborate with investigators from the ACTG 5142 study, which randomized treatment naove HIV patients to three different treatment strategies. The A5142 study will allow us to evaluate the significance of rising residual viremia and its association with the emergence of ART resistance in an HIV-1 treatment naove population failing recommended ART strategies. We will use the Single Copy Assay, a highly sensitive viral load assay, to determine whether rising HIV-1 residual viremia is prevalent in patients with subsequent virologic failure. Emergence of resistance will be assessed by standard genotyping and by the novel Single Genome Sequencing method, to allow detection of emergence of minor resistant variants present as low as 5% of the viral population. This proposal will improve our understanding of the dynamics of viral rebound, and may identify an earlier window for clinical interventions to preserve therapeutic options. This innovative, high impact project has the potential to fundamentally alter monitoring of patients on antiretroviral therapy. PUBLIC HEALTH RELEVANCE: This application investigates whether an increase in low-level residual viremia (HIV-1 RNA) in patients on antiretroviral therapy is an early indicator of impending virologic failure and is associated with the emergence of minor or major populations of drug-resistant HIV-1 variants.